Drug discovery is a long, multiple step process involving identification of specific disease targets, development of an assay based on a specific target, validation of the assay, optimization and automation of the assay to produce a screen, high throughput screening of compound libraries using the assay to identify “hits”, hit validation, and hit compound optimization. The output of this process is a lead compound that goes into pre-clinical studies and, if validated, eventually into clinical trials. In this process, the screening phase is distinct from the assay development phases, and involves testing compound efficacy in living biological systems.
Historically, drug discovery is a slow and costly process, spanning numerous years and consuming hundreds of millions of dollars per drug created. Developments in the areas of genomics, proteomics, and high throughput screening have resulted in increased capacity and efficiency in the areas of target identification, structure-function predictions, and volume of compounds screened. Significant advances in automated DNA sequencing, PCR application, positional cloning, hybridization arrays, and bioinformatics have greatly increased the number of genes (and gene fragments) encoding potential drug screening targets. However, the basic scheme for drug screening remains the same.
The next level of biological complexity is the cell, and sophisticated automated methods for cell-based screening based on imaging of fluorescent reporter molecules in cells have recently been developed. (See, for example, U.S. Pat. Nos. 5,989,835 and 6,103,479, as well as published PCT application Nos. WO 98/38490, WO 00/03246, WO 00/17643, WO 00/26408, WO 00/50872, WO/00/70342, WO 00/17624, and WO/00/60356.) The process of implementing such cell-based assays is also referred to as high content screening (“HCS”), and addresses a need for more detailed information about the temporal-spatial dynamics of cell constituents and processes, and how they are affected by potential drug candidates.
HCS automates the extraction of multicolor luminescence information derived from specific luminescence-based reagents incorporated into cells (Giuliano and Taylor (1995), Curr. Op. Cell Biol. 7:4; Giuliano et al. (1995) Ann. Rev. Biophys. Biomol. Struct. 24:405). Cells are analyzed using an optical system that can measure spatial, as well as temporal dynamics. (Farkas et al. (993) Ann. Rev. Physiol. 55:785; Giuliano et al. (1990) In Optical Microscopy for Biology. B. Herman and K. Jacobson (eds.), pp. 543–557. Wiley-Liss, New York; Hahn et al (1992) Nature 359:736; Waggoner et al. (1996) Hum. Pathol. 27:494). The concept is to treat each cell as a “well” that has spatial and temporal information on the activities of the labeled constituents.
HCS can be performed on living or fixed cells, using a variety of labeled reporter molecules, such as antibodies, biological ligands, nucleic acid hybridization probes, and multicolor luminescent indicators and “biosensors.” The choice of fixed or live cell screens depends on the specific cell-based assay required.
The results obtained from HCS provide more relevant information about a drug candidate's potential effect on cells than is available from genomic or proteomic methods, and can dramatically reduce costs in animal testing, while increasing the speed of new drug development.
While HCS can be combined on a single platform with high throughput screening (HTS) (see, for example, U.S. Pat. Nos. 5,989,835 and 6,103,479, as well as published PCT application no. WO 98/38490), methods that further increase the throughput capabilities of high content cell-based drug screening would be of great value to the art. The average “hit” rate (i.e.: detection of a positive response) in most viable high content cell-based screens ranges from 0.1% to 1.0% of compounds screened. Therefore, the vast majority of wells screened in a microplate format yield a negative response, necessitating the screening of a large number of wells to detect a response of interest, which significantly impacts the capacity requirements of such high content cell-based screening assays. Thus, methods that increase the capacity to provide high content information on the effect of a test compound on cellular events of interest, while maintaining the “hit” rate, would provide a tremendous increase in the utility of high content cell based screens.